Genome damage and defective repair are etiologically linked to Fused in Sarcoma (FUS)-associated amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). However, the underlying mechanisms remain enigmatic, which is a roadblock for exploiting genome repair-targeted therapies for ALS/FTD. Our recent publication (Wang et al, Nature Communications, 2018) identified defects in DNA nick ligation and oxidative damage repair in a subset of ALS patients, caused by mutations in the RNA/DNA-binding protein FUS. In healthy neurons, FUS protects the genome by facilitating PARP1-dependent recruitment of XRCC1/DNA Ligase III? (LigIII) to oxidized genome sites and activates LigIII via direct interaction. We discovered that FUS toxicity caused significantly decreased recruitment of XRCC1/LigIII to DNA strand breaks. DNA ligation defects in ALS patient-derived iPSC lines carrying FUS mutations and in subsequently generated motor neurons were rescued by CRISPR/Cas9-mediated mutation correction. Moreover, our follow-up studies showed substantially reduced auto and total PARylation activity of PARP-1 both in vitro and in cell, after loss of FUS or mutant expression, which in addition to regulating LigIII/XRCC1 recruitment at damage sites, could impact neuronal energy metabolism by uncoupling NAD+/NADH levels and stress granule dynamics in motor neurons. Collectively these events may provide a recipe for neurodegeneration. These findings that uncovered a new pathway of defective DNA ligation and PARP-1 functions in FUS-linked ALS-FTD, raised three key questions that need to be investigated to understand their implications in neuronal death and to develop a comprehensive strategy of PARylation and LigIII targeted interventions for ameliorating FUS-associated ALS-FTD. These questions are: (1) How does FUS affect PARP-1's PARylation activity and what is its impact on genome maintenance and energy metabolism? (2) What is the effect of FUS-mediated LigIII inhibition on the mitochondrial genome and its functions? This is important as LigIII is the only DNA ligase for both replication and repair in mitochondria, and both FUS and PARP-1 localize in mitochondria. (3) What is the effect of FUS pathology on microhomology-mediated alternative end-joining (MMEJ) pathway of DNA double strand break repair, which involves LigIII, XRCC1 and PARP-1?. This project, will utilize human patient-derived iPSC lines harboring FUS mutations, their isogenic controls with mutation correction by CRISPR/Cas9 knock-in strategy, a transgenic FUS-?NLS mouse model and human ALS, FTD patient spinal cord/brain tissue, to test our novel hypothesis that FUS pathology-mediated DNA ligation defects via reduced PARylation inhibits oxidative genome damage repair and promotes neurodegeneration. We will further show that rescuing Ligase and PARP functions are promising avenues for neuroprotection. Our studies investigating the previously unexplored link between altered FUS-PARP-1-LigIII axis and ALS-FTD are both technically and conceptually innovative, have important immediate and long term goals and will strongly impact translational ALS-FTD research.